Biliverdin reductase (“BVR”) catalyzes reduction of the γ-meso bridge of biliverdin, an open tetrapyrrole, to produce bilirubin (Singleton et al., J. Biol. Chem. 240: 47890-4789 (1965); Tenhunen et al., Biochemistry 9:298-303 (1970); Colleran et al., Biochem J. 119:16P-19P (1970); Kutty et al., J. Biol. Chem. 256:3956-3962 (1981); Buldain et al., Eur. J. Biochem. 156:179-184 (1986); Noguchi et al., Biochem J. 86:833-839 (1989)). In mammals, the oxidative cleavage of heme is catalyzed by the heme oxygenase system (Maines, Ann. Rev. Pharmacol. Toxicol. 37:517-554 (1997)). Because open tetrapyrroles are generally believed to be devoid of biological functions, the enzymes that catalyze their formation have not traditionally been in the main stream of research activity. In plants, however, biliverdin analogues, phytochromobilins, function in photoregulatory capacity (Terry et al., J. Biol. Chem. 266:22215-22221 (1991); Cornejo et al., J. Biol. Chem. 267:14790-14798 (1992)). Molecular cloning and biochemical analyses have shown that the enzyme, which in human is a 296 residue polypeptide, is highly conserved both at its primary structure and at its unique catalytic properties (Fakhrai et al., J. Biol. Chem. 267:4023-4029 (1992); McCoubrey et al., Eur. J Biochem. 222:597-603 (1994); McCoubrey et al., Gene 160:235-240 (1995); Maines et al., Eur. J. Biochem. 235:372-381 (1996)). BVR is the only enzyme described to date with dual pH/dual adenine nucleotide cofactor requirements (Kutty et al., J. Biol. Chem. 256:3956-3962 (1981); Fakhrai et al., J. Biol. Chem. 267:4023-4029 (1992); Maines et al., Eur. J. Biochem. 235:372-381 (1996); Huang et al., J. Biol. Chem. 264:7844-7849 (1989)). The reductase uses NADH in the acidic range (optimum range ˜pH 6.0-6.7), whereas NADPH is utilized in the basic range (optimum range ˜pH 8.5-8.7). BVR, which is a zinc metalloprotein (Maines et al., Eur. J. Biochem. 235:372-381 (1996)), possesses a HCX10CH or HCX10CC motif in the carboxy terminal third of the protein, which is similar to the zinc binding motif of protein kinase C (Hubbard et al., Science 254:1776-1779 (1991)) and may be the site of interaction of BVR with zinc.
BVR was previously thought to be simply a house-keeping enzyme found in most mammalian cells in excess of, or in disproportionate levels to, heme oxygenase isozymes (Ewing et al., J. Neurochem. 61:1015-1023 (1993)). Yet it has the above-noted noted unique and uncommon properties. Examination of the primary structure of human BVR, which recently became available (Maines et al., Eur. J. Biochem. 235:372-381 (1996)), revealed the presence of consensus sequences that are conserved in protein kinases, the most notable one being the GXGXXG motif near the N terminus of the protein that is found invariably in all kinases (Kamps et al., Nature 310:589-592 (1984); Hunter et al., Ann. Rev. Biochem. 54:897-930 (1985); Schlessinger, Trend. Biochem. Sci. 13:443-447 (1988); Hanks et al., Science 241:42-52 (1988); Yarden et al., Annu. Rev. Biochem. 57:443-478 (1988); Hanks et al., Methods Enzymol. 200:38-62 (1991)). A valine residue is present in BVR just 2 positions downstream from the last glycine of this motif. A valine residue is invariant at the corresponding position, as in BVR, in the family of kinases that phosphorylate G-protein coupled receptors (Garcia-Bustos et al., Biochim. Biophys. ACTA 1071:83-101 (1991)). Database search results also identified additional similarities with PKGs, including a cluster of charged residues (KRNR) in the carboxy terminus of BVR. Such clusters are a characteristic of the nuclear localization signal (“NLS”) (Garcia-Bustos et al., Biochim. Biophys. ACTA 1071:83-101 (1991)).
Although BVR has previously been identified as exhibiting protective effects against oxidative stress and as sharing characteristics with known kinases (see U.S. patent application Ser. No. 09/606,129 to Maines, filed Jun. 28, 2000), it is unclear the extent to which BVR is implicated in cellular repair mechanisms. The present invention is directed to overcoming the above-identified deficiencies in the art.